This proposal is to continue studies on Ca2+-mediated synaptic transmitter release from cultured amacrine cells from chicken retina. Conventional patch-clamp techniques together with fluorometric Ca2+-imaging measurements will be used. Four overlapping groups of experiments are planned. The first set of experiments is intended to discover what concentration of cytosolic Ca2+ is required for quantal release of transmitter from amacrine cells. These experiments are designed to test the supposition, based on previous studies that Ca2+ concentrations much lower than generally supposed are sufficient for quantal release. A second set of experiments will test the idea that miniature currents at central synapses are, unlike those at the neuromuscular junction, responses to the simultaneous release of several vesicles, coordinated by local Ca2+ domains. This idea involves eliciting transmitter release in the absence of Ca2+ domains and using Ca2+-imaging to look for the existence of domains. A third set of studies will continue investigations of the mechanisms by which Ca2+ is removed from synapses and transmission is turned off. Outstanding questions in this regard are the stoichiometry of Na+/Ca2+ exchange and the relative importance of mitochondria and endoplasmic reticulum in Ca2+ uptake. Lastly, experiments will investigate the mechanism(s) by which the retinal peptide, neurotensin, found in amacrine cells elicits Ca2+ oscillations and transmitter release.